ESCRIPTION (provided by applicant): Transforming growth factor-beta (TGF-beta) and members of the TGF-beta family play key roles in the regulation of cell proliferation and differentiation, and in normal development and tumorigenesis. They signal through heteromeric complexes of two types of cell surface, transmembrane serine-threonine kinases, resulting in phosphorylation and consequent activation of Smads, which then act as signaling effectors in the regulation of TGF-beta-induced gene expression. Extensive research during the last few years has revealed a general model for how receptor-activated Smads activate transcription through cooperation with sequence-specific transcription factors at the promoter DNA. In contrast, little is known about the mechanisms that result in downregulation of gene expression in response to TGF-beta. We now propose to continue our ongoing research on the regulation of gene expression in response to TGF-beta. In Aim 1, we will study the mechanism of TGF-beta/Smad3-mediated repression of transcription by MyoD/MEF2 and CBFA1, which naturally drive differentiation in myoblasts and osteoblasts, respectively. The molecular characterization of these two model systems for TGF-beta/Smad3-mediated repression should provide insight into general features and may lead to a general model for the mechanism of transcription repression by TGF-beta family members through Smads. Furthermore, the study of the regulation of CBFA1 function will allow us to characterize the roles of the CBFAl-binding DNA sequence and the cell context (mesenchymal versus epithelial) in TGF-beta-mediated repression versus activation of transcription. Aim 2 will focus on the crosstalk between IRF-3/7 and Smad signaling and the role of IRF-3/7 in TGF-b/Smad signaling. IRF-3 and IRF-7 are two closely related members of the "interferon regulatory factor" (IRF) family of transcription factors. Viral infection activates IRF-3/7 and induces IRF-7 expression, which leads to type I interferon and RANTES expression. We will characterize the crosstalk between IRF-3/7 and TGF-beta/Smad signaling at the IRF-3/7-responsive interferon b promoter, and conversely evaluate the effect of IRF-3/7 on Smad-mediated transcription. We will also evaluate the role of TGF-beta/Smad signaling in the antiviral induction of interferon, driven by IRF-3/7, and evaluate a possible direct role of IRF-3/7 in TGF-beta signaling. Mutation analyses of IRF-3/7, based on the three-dimensional IRF-3, structure will assist in these investigations.